calyculin-a and Alzheimer-Disease

Berberine is a primary component of the most functional extracts of Coptidis rhizome used in traditional Chinese medicine for centuries. Recent reports indicate that Berberine has the potential to prevent and treat Alzheimer's disease (AD). The previous studies reported that Calyculin A (CA) impaired the axonal transport in neuroblastoma-2a (N2a) cells. Berberine attenuated tau hyperphosphorylation and cytotoxicity induced by CA. Our study aimed at investigating the effects of Berberine on the axonal transport impairment induced by CA in N2a cells. The results showed that Berberine could protect the cell from CA -induced toxicity in metabolism and viability, as well as hyperphosphorylation of tau and neurofilaments (NFs). Furthermore, Berberine could reverse CA-induced axonal transport impairment significantly. Berberine also partially reversed the phosphorylation of the catalytic subunit of PP-2A at Tyrosine 307, a crucial site negatively regulating the activity of PP-2A, and reduced the levels of malondialdehyde and the activity of superoxide dismutase, markers of oxidative stress, induced by CA. The present work for the first time demonstrates that Berberine may play a role in protecting against CA-induced axonal transport impairment by modulating the activity of PP-2A and oxidative stress. Our findings also suggest that Berberine may be a potential therapeutic drug for AD.

Topics: Alzheimer Disease; Animals; Axonal Transport; Axons; Berberine; Cell Line, Tumor; Cell Shape; Malondialdehyde; Marine Toxins; Mice; Neurons; Oxazoles; Oxidative Stress; Phosphorylase b; Phosphorylation; Superoxide Dismutase; tau Proteins

Abnormal phosphorylation and aggregation of the microtubule-associated protein Tau are hallmarks of various neurodegenerative diseases, such as Alzheimer disease. Molecular mechanisms that regulate Tau phosphorylation are complex and currently incompletely understood. We have developed a novel live cell reporter system based on protein-fragment complementation assay to study dynamic changes in Tau phosphorylation status. In this assay, fusion proteins of Tau and Pin1 (peptidyl-prolyl cis-trans-isomerase 1) carrying complementary fragments of a luciferase protein serve as a sensor of altered protein-protein interaction between Tau and Pin1, a critical regulator of Tau dephosphorylation at several disease-associated proline-directed phosphorylation sites. Using this system, we identified several structurally distinct GABA(A) receptor modulators as novel regulators of Tau phosphorylation in a chemical library screen. GABA(A) receptor activation promoted specific phosphorylation of Tau at the AT8 epitope (Ser-199/Ser-202/Thr-205) in cultures of mature cortical neurons. Increased Tau phosphorylation by GABA(A) receptor activity was associated with reduced Tau binding to protein phosphatase 2A and was dependent on Cdk5 but not GSK3β kinase activity.

Topics: Alzheimer Disease; Animals; Cell Line, Tumor; Cyclin-Dependent Kinase 5; Cytoskeleton; Enzyme Inhibitors; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Humans; Marine Toxins; Mice; Naphthoquinones; Nerve Degeneration; Neuroblastoma; NIMA-Interacting Peptidylprolyl Isomerase; Oxazoles; Peptidylprolyl Isomerase; Phosphorylation; Protein Kinase Inhibitors; Protein Phosphatase 2; Purines; Rats; Receptors, GABA-A; Roscovitine; tau Proteins; Tauopathies

We have reported recently that inhibition of protein phosphatase (PP)-2A and PP-1 by calyculin A, a specific inhibitor of PP-2A and PP-1, induced Alzheimer-like hyperphosphorylation of tau and spatial memory retention impairment. In this study, we tested the in vivo effects of melatonin on these Alzheimer-like changes. We found that administration of melatonin intraperitoneally for 9 consecutive days before injection of calyculin A could prevent calyculin A-induced synaptophysin loss, memory retention deficits, as well as hyperphosphorylation of tau and neurofilaments. Furthermore, melatonin partially reversed the phosphorylation of the catalytic subunit of PP-2A at Tyrosine 307 (Y307), a crucial site negatively regulating the activity of PP-2A, and reduced the levels of malondialdehyde, a marker of oxidative stress, induced by calyculin A. These results suggest that melatonin could serve as a potential therapeutic agent for preventing Alzheimer-like pathological changes and behavioral abnormality via modulating the activity of PP-2A and oxidative stress.

Topics: Alzheimer Disease; Animals; Antioxidants; Behavior, Animal; Disease Models, Animal; Male; Malondialdehyde; Marine Toxins; Melatonin; Memory Disorders; Oxazoles; Oxidative Stress; Phosphorylation; Protein Phosphatase 2; Rats; Rats, Sprague-Dawley; tau Proteins

Human fibroblast cell culture systems have been used to model both molecular events associated with the aging process and the biochemical anomalies found in the aging-associated neurodegenerative disorder Alzheimer's disease (AD). We demonstrate modulation of bradykinin (BK) B2 receptors that results in Intermediate (I, Kd 2.5-5 nM) and Low (L, Kd 44 nM) receptor affinity states in two cellular model systems that target aging and aging-associated disorders: the human lung fibroblast cell line WI-38 model for cellular aging and a skin fibroblast cell line from a patient with early onset familial Alzheimer's disease. In both cellular models the generation of I and L BK B2 receptors is extremely rapid, occurring within 1 min of activation of protein kinase C (PKC) by phorbol ester. Blocking phosphoprotein phosphatase activity further augments the cellular content of I and L receptors in the Alzheimer's skin fibroblast cell line. These two lines of evidence suggest that a phosphorylation cascade modifying the receptors is responsible for the I and L states. The I and L receptors remain biologically active and enhance cellular responsiveness to elevated levels of BK that are found in tissue injury, one of the major risk factors for development of Alzheimer's disease. The Alzheimer's disease skin fibroblast cell line presents a cellular environment highly enriched in the amyloid Abeta1-42 peptide that is the hallmark of Alzheimer's plaque lesions in the brain. This Abeta-rich environment may serve to foster the signal transduction mechanism that generates I and L BK B2 receptors.

Topics: Aging; Alzheimer Disease; Blotting, Western; Cell Line; Electrophoresis, Polyacrylamide Gel; Fibroblasts; Humans; Lung; Male; Marine Toxins; Oxazoles; Phorbol Esters; Phosphoprotein Phosphatases; Phosphorylation; Protein Kinase C; Receptor, Bradykinin B2; Receptors, Bradykinin; Skin

Aberrant metabolism of the Alzheimer amyloid precursor protein (APP) or its amyloidogenic A beta fragment is thought to be centrally involved in Alzheimer's disease. Nonamyloidogenic processing of APP involves its cleavage within the A beta domain by a protease, termed alpha-secretase, and release of the large extracellular domain, termed APPS. Secretion of APPS can be stimulated by phorbol esters, activators of protein kinase C, with concurrent inhibition of A beta production. While the role of protein kinases of APP metabolism has been investigated, considerably less effort has been devoted to elucidating the role played by protein phosphatases. Okadaic acid, a protein phosphatase inhibitor, has been shown to stimulate secretion of APPS, but the identity of the phosphatase involved has not been investigated.. The secretion of APPS from COS-1 cells was measured in the absence or presence of various doses of serine/threonine-specific phosphatase inhibitors. Quantitation of the derived IC50 values was used to determine the identity of the phosphatase involved in the control of APP secretion.. The availability of protein phosphatase inhibitors with different relative potencies against the different types of serine/threonine-specific protein phosphatase allowed us to examine which of the four known types of protein phosphatase might be involved in the regulation of APP secretion. Both okadaic acid and calyculin A stimulated the secretion of APP from COS-1 cells in a dose-dependent manner. The half-maximal dose for stimulation of APP secretion was approximately 100-fold higher with okadaic acid than with calyculin A.. The nearly 100-fold difference in the observed IC50 values for okadaic acid and calyculin A implicates a type 1 protein phosphatase in the control of APPS production. Protein phosphatase 1 (PP1) is known to be highly expressed in adult mammalian brain, both in neurons and glia. The identification of a specific phosphatase type in the control of APP secretion opens new avenues to the development of rational therapeutic intervention strategies aimed at the prevention and/or treatment of Alzheimer's Disease.

Topics: Aged; Alzheimer Disease; Amyloid; Amyloid Precursor Protein Secretases; Aspartic Acid Endopeptidases; Cantharidin; Cell Line; Endopeptidases; Enzyme Inhibitors; Ethers, Cyclic; Humans; Immunoblotting; Marine Toxins; Okadaic Acid; Oxazoles; Phorbol 12,13-Dibutyrate; Phosphoprotein Phosphatases; Prion Proteins; Prions; Protein Phosphatase 1; Protein Precursors